Hepatitis is a general term meaning ‘inflammation of the liver’ and has a number of causes. Viral causes are among the most common, and may be caused by hepatitis A, B, C, D or E virus. Hepatitis B virus (HBV) in particular is a serious and common infectious disease of the liver, affecting millions of people throughout the world.
HBV is a hepatotrophic DNA virus belonging to the Hepadnaviridae. The full-length of the viral genome is about 3.2 kb, and it has four open reading frames (ORFs) including surface antigen (the “S gene”), core antigen (the “C gene”), DNA polymerase (the “P gene”) and a gene of undetermined function referred to as the “X gene”.
More than 2,000 million people alive today have been infected with HBV at some time in their lives and of these about 350 million remain chronically infected and become carriers of the virus. HBV infection can cause acute and chronic type B hepatitis, and may eventually lead to the development of chronic hepatic insufficiency, cirrhosis, and hepatocellular carcinoma. In addition, HBV carriers can transmit the disease for many years.
HBV is transmitted by percutaneous or parenteral contact with infected bodily fluids or blood. The most common route of infection is via vertical transmission from mother to her baby, and in adults through sexual intercourse or shared intravenous needles or ear-piercing equipment. Many cases of acute HBV infection occur however without a traceable route of infection.
Persons with chronic HBV infection (“carriers”—worldwide about 350-400 million people) have a 12-300× higher risk of developing hepatocellular carcinoma than non-carriers and globally HBV causes 60-80% of the world's primary liver cancers. Every year about 25% of the over 4 million acute clinical cases (i.e. 1 million people worldwide) die from chronic active hepatitis, cirrhosis or HBV-induced liver cancer. As a consequence, HBV ranks second only to tobacco as a known human carcinogen.
Although vaccines against HBV has been widely used for several decades, the HBV prevalence rate in the population still remains high. Current therapies for chronic HBV infection have only limited inhibitory effects on viral gene expression and replication in the majority of chronically infected patients. Lamivudine for example suppresses HBV replication in carriers, but the effect is reversible if therapy is stopped. Moreover, a major limitation of chronic Lamivudine therapy is the development of viral resistance, which typically develops after 6 months of treatment. Resistance is usually associated with mutations in the highly conserved catalytic region of the HBV polymerase gene.
For these reasons, there remains a need for a new therapeutic agent to treat HBV infection. This invention is directed to an RNA interference (RNAi) agent and the use of that RNAi agent to treat hepatitis B infection in individuals.
The RNAi pathway is initiated by the enzyme Dicer, which cleaves double-stranded RNA (dsRNA) molecules into short fragments (commonly referred to as siRNAs) of ˜20-25 nucleotides. One of the two strands of each fragment, known as the guide strand or active strand, is then incorporated into the RNA-induced silencing complex (RISC) through binding to a member of the argonaute protein family. After integration into the RISC, the guide strand base-pairs with its target mRNA and is thought to either inhibit a target by inhibiting translation (by stalling the translational machinery) and/or inducing cleavage of the mRNA, thereby preventing it from being used as a translation template.
While the fragments produced by Dicer are double-stranded, only the guide strand, directs gene silencing. The other anti-guide strand referred to more commonly as a passenger strand, carrier strand or * strand is frequently degraded during RISC activation (Gregory R, Chendrimada T, Cooch N, Shiekhattar R (2005). “Human RISC couples microRNA biogenesis and posttranscriptional gene silencing”. Cell 123 (4): 631-40). RISC assembly is thought to be governed by an enzyme that selects which strand of a dsRNA Dicer product is loaded into RISC. This strand is usually the one whose 5′ end is less tightly paired to its complement, and there also appears to be a clear bias for A, and to a lesser extent U, at the 5′ position to facilitate binding to some argonaute proteins (Schwarz D S, Hutvagner G, Du T, Xu Z, Aronin N, Zamore P D (2003). “Asymmetry in the assembly of the RNAi enzyme complex”. Cell 115 (2): Frank F, Sonenberg N, Nagar (2010) “Structural basis for 5′-nucleotide base-specific recognition of guide RNA by human AGO2”. Nature. 465(7299):818-22).
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